Telomeres are found at the end of linear chromosomes and consist of short repetitive sequences essential for the maintenance of normal chromosome structure and function (Wellinger & Sen, 1997). With each cell division, telomeres shorten due to the inability of DNA polymerases to replicate the ends of linear DNA molecules. However, telomere erosion is counteracted by the activity of the enzyme telomerase, a ribonuclear protein with reverse transcriptase activity, which adds telomeric repeats to the chromosomal termini (Morin, 1997; Nakamura et al., 1997). The genes for the human, (hTR), and mouse, (terc), RNA components have recently been cloned, as has the human protein component, (hTRT) (Blasco et al., 1995; Feng et al., 1995; Nakamura et al., 1997; Soder et al., 1997b; Soder et al., 1997c). Whilst telomerase expression is detectable in normal embryonic tissues and germline stem cells, telomerase expression is repressed in most normal postnatal somatic cells (Blasco et al., 1995; Feng et al., 1995; Prowse & Greider, 1995; Soder et al., 1997a; Wright et al., 1996). The lack of telomerase expression may be the major reason for the progressive loss of telomeric sequences in somatic cells, which is considered to be one regulatory mechanism which monitors the number of times a cell divides before entering replicative senescence (Campisi, 1997). However, although telomerase appears to be stringently repressed in normal somatic tissues, there is substantial evidence to suggest that telomerase is expressed in the majority of human cancers and contributing to the immortal phenotype through the maintenance of telomere integrity (Holt et al., 1997; Kim, 1997; Shay & Bacchetti, 1997).
The regulation of telomerase expression is a complex issue including transcriptional activity of the telomerase RNA and protein component genes, interaction of telomerase with other telomere associated proteins and post-translational modification of the enzyme complex. However, at present there are few studies which directly address the mechanisms regulating telomerase expression in normal and cancer cells (Bodnar et al., 1996; Broccoli et al., 1997; Li et al., 1997; Mandal & Kumar, 1997; Morin, 1997; Nakamura et al., 1997; Soder et al., 1997a).
Studies aimed at relating genome stability to human cellular senescence have recently placed considerable emphasis on telomerase expression as a central unifying mechanism underlying the immortal phenotype of many cancers (Breslow et al., 1997). The absence of telomerase activity from normal somatic cells has led to the proposal that telomere shortening may be a molecular clock which contributes to the onset of cellular senescence in normal cells (Harley & Villeponteau, 1995; Holt et al., 1996). Conversely, the reactivation or expression of telomerase may be a major mechanism by which cancer overcome normal cellular senescence (Kim et al., 1994; Parkinson et al., 1997). Information on telomerase activity in tumours almost exclusively derives from the in vitro telomere repeat amplification protocol, (TRAP), and these have shown that telomerase activity may be present in greater than 80% of tumour biopsies yet absent or reduced in normal somatic tissue (Breslow et al., 1997; Kim et al., 1994; Raymond et al., 1996; Shay & Wright, 1996). However, TRAP assay alone will not reveal the true complexities of telomerase regulation, and it is generally recognised that a number of molecular approaches will be required to understand telomere length regulation and telomerase activity (Breslow et al., 1997; Holt et al., 1996; Lundblad & Wright, 1996; Parkinson et al., 1997; Raymond et al., 1996; Soder et al., 1997b). Recently, the present inventors and others have introduced a number of more direct in situ approaches to study the telomerase RNA gene, (hTR) and its expression in tumours (Soder et al., 1997; Yashima et al., 1997b).
U.S. Pat. No. 5,583,016 (Geron Corp.) Discloses a 2.4 kb sequence of the Telomerase RNA gene. However, there is no disclosure of the promoter elements or provision of functional evidence to show the promoter is active.